The proposed study will examine the mechanisms underlying inward rectification in cloned potassium channels, utilizing a combination of molecular biological and electrophysiological techniques. In preliminary experiments, two novel inward rectifying potassium channel cDNAs have been cloned and characterized, and a soluble cytoplasmic factor (IRF) responsible for intrinsic rectification has been isolated. Based on background and preliminary data, the hypothesis is developed that voltage- dependent rectification and gating of inward rectifier potassium channels involve fundamentally similar processes and structural elements to those responsible for the gating of voltage-dependent potassium channels. In order to extend preliminary data and examine the above hypothesis, five experimental series aimed of experiments are proposed to address the following questions: (1) What structural features underlie natural diversity of inward rectification? (2) What is the chemical identity of IRF and how does it produce inward rectification? (3) What specific structural elements are involved in Mg2+ block of potassium channels? (4) Is the M0 domain a voltage sensor responsible for hyperpolarization deactivation of inward rectifier channels? (5) What mechanisms are involved in physiological modulation of inward rectification? The results of the proposed experiments, answering the above questions, will provide detailed insight into the fundamental mechanism of inward rectification, a critical determinant of the functional diversity of potassium channels. Inward rectification is essential for regulation of cell excitability and potassium homeostasis in cardiac, brain and other tissues. The work will therefore provide information that will ultimately underlie the development of rational therapies for the treatment of cardiac arrhythmias, epilepsy and other disorders of cell excitability.